Studies in Project 11 address the hypothesis that neural cell death after neonatal hypoxia/ischemia (H/l) arises from both acute impairment in mitochondrial energy metabolism and prolonged metabolic alterations that leave the brain vulnerable to secondary insults including damage from hyperoxic resuscitation. Our studies show that H/l leads to acutely impaired mitochondrial respiration and decreased activity of pyruvate dehydrogenase complex (PDHC) and a-ketoglutarate dehydrogenase (KGDH), loss of aralar1 (the mitochondrial aspartate-glutamate carrier), and long term impairment of metabolism and GABA synthesis in brain. Aralar1, which links the loss of NAD*, energy failure and decreased synthesis of the neuronal integrity marker N-acetylaspartate (NAA), is an important new target for neuroprotection. Our data provide evidence that treatment of pups after H/l injury with acetyl-L-carnitine (ALCAR) protects brain proteins and metabolism. We hypothesize that optimal neuroprotection following H/l injury can be achieved by preserving cerebral energy metabolism, key proteins and GABA synthesis through the post-injury administration of ALCAR. sulforaphane. and/or estradiol; therapeutic agents that protect by different mechanisms. The following Specific Aims will test these hypotheses: 1) Determine if both early and delayed alterations in mitochondrial proteins involved in energy metabolism after H/l are alleviated by normoxic resuscitation and treatment with ALCAR, SFP, estradiol or combined therapy; 2) Determine if the early and long-term in vivo brain metabolic and structural changes (detectable by serial spectroscopy and imaging) after H/l are alleviated by ALCAR; 3) Determine the long-term effects of H/l on neuronal and glial pathways of metabolism and synthesis of glutamate and GABA by C-NMR spectroscopy and determine the cell specific efficacy of ALCAR for protecting key pathways of metabolism. A powerful combination of biochemical methods, in vivo 31 P and[1]'H MRS. T2 and diffusion weighted imaging (DWI), diffusion tensor imaging (DTI), and [13]C-NMR, will be used to test these hypotheses. Results are expected to reveal unique insights into the timing, targets and mechanisms of injury in both neuronal and glial metabolic pathways after H/l.